Method for transferring heterogeneous liquids in microchannels without the occurrence of mixing

ABSTRACT

In the method for mixing-free transport of homogeneous liquids in microchannels a heterogeneous liquid flow ( 14 ) is autonomously divided into portions, transported over a distance and then autonomously recombined. When the flow is divided into portions, individual volumes of a second liquid ( 16 ), which may be but must not be homogeneous, are introduced between volume fractions lying one behind the other of the heterogeneous liquid flow ( 14 ). While the volume fractions of the heterogeneous liquid ( 14 ) that lye close to one another would mix with each other without this measure being taken, separation of the individual volume fractions allows these volume fractions, and thus the heterogeneous liquid ( 14 ), to be transported over large distances without any mixing occurring. Treatment of the volume fractions of the heterogeneous liquid ( 14 ) requires separation of the liquid flow ( 12 ) of alternating volume fractions of the two liquids ( 14,16 ) such that again two flows are produced, one of which being the heterogeneous starting liquid and the other being the second liquid. Since both processes can take place autonomously by self-organization of the phases, an overall solution for the mixing-free transport of a heterogeneous liquid without employment of expensive fractionating apparatus can be achieved.

The invention relates to a method for mixing-free transport ofheterogeneous liquids in microchannels. In particular, the inventionrelates to the mixing-free transport of a heterogeneous liquid betweentwo locations in a channel system.

TECHNICAL FIELD OF APPLICATION

Heterogeneous liquids, i. e. liquids comprising concentration gradientsof molecules or microparticles (e. g. beads), are produced in a varietyof synthetic or analytical tasks in the chemical sector. Above all inconnection with flow reactors or separation processes, such aschromatography and gel electrophoresis (PAGE) in the field ofbiotechnology, but also in the field of combinatorial chemistry or in“Lap on a chip” applications, heterogeneous liquids must be transportedin a mixing-free manner from the location of their production to thelocation of further treatment (e. g. analysis, fractioning or othertreatment steps).

STATE OF THE ART

In conventional analysis processes, such as chromatography orelectrophoresis, heterogeneously fractioned solutions suffer, duringtheir transport within capillary systems, from a strong longitudinalmixing, which strongly reduces their dissolution capacity. This is dueto the development of the well-known parabolic velocity profile whichcauses the flow in the middle of the channel or the capillary to movefaster than at the edge. Consequently, a lateral diffusion causes, overshort distances, the longitudinal mixing in the flow (G. TaylorConvection, cf. “Dispersion of soluble matter in solvent flowing slowlythrough a tube, Proc. Roy. Soc., London 219A (1953) 186-203). Although adecrease in the tube diameter reduces the overall mixing, it increasesthe hydrodynamic resistance and causes larger problems due to walleffects and adsorption. Recent research has shown that a targetedlateral mixing caused by microstructured components reduces this effectto a certain degree (cf. M. J. Clifton “Continuous flow electrophoresisin the Taylor regime”, J. Chromatography A 757 (1997) 193-202). However,this so-called “Taylor dispersion” and the wall adsorption remainlimiting factors for a continuous single-phase sample transport incapillaries.

Spontaneous phase separations of immiscible liquids are known (cf. K.-V.Schubert and E. W. Kaler “Nonionic microemulsions”, Ber.Bunsengesellschaft 100 (1996) 190-205). Thus, for example, equilibriumoil and water form a two-phase system. The production of droplets fromimmiscible phases within microchannels has also been analysed. Incontrast, the utilization of droplets from immiscible phases for themixing-free transport of microscopic sample volumes has not yet beendescribed. Further, the control of non-equilibrium phase formationthrough surface, geometry and flow velocity effects has been the subjectof little research so far (cf. L. M. Grant and W. A. Ducker “Effect ofSubstrate Hydrophobicity on Surface-Aggregate Geometry”, J. Phys. Chem.B101 (1997) 5337-5345). Surface properties such as hydrophobicity andhydrophilicity are utilized in targeted positioning of droplets at opensurfaces and keeping them at defined locations (e. g. in water-airsystems). The formation of droplet chains in air-water systems andoil-water systems is known (cf. “Droplet formation in a microchannelnetwork”, Takasi Nisiako, Toru Torii, Toshiro Higuchi, Lab on a Chip,Vol. 2, No. 1 (2002) and “Dynamic Pattern Formation in aVesicle-Generating Microfluidic Device”, Todd Thorsen, Richard W.Roberts, Frances H. Arnold and Stephen R. Quake, Physical ReviewLetters, Vol. 86, No. 18 (2001)), but not their utilization inconnection with autonomous separation of droplets for targeted transportof heterogeneous samples. Water-air systems have the further drawbackthat they are adapted to expand under changing pressure conditions andare not precisely controllable.

OBJECT ACHIEVED BY THE INVENTION

The invention describes a device and a method for “digitally” encodingand decoding heterogeneous liquids in discrete droplets, which isrendered possible by mixing-free transport within capillaries and/ormicrochannels.

Solution

According to the invention, this objective is achieved with a method formixing-free transport of a heterogeneous liquid, i. e. a liquid which,as seen in the direction of flow, comprises a inhomogeneous component,in microchannels, wherein in the method

-   -   a first heterogeneous liquid flows through a first channel and,        at a first connecting location, meets a second channel carrying        a second liquid (hereinafter referred to as separating liquid,        wherein this second liquid may be homogeneous or inhomogeneous),        wherein the two liquids are selected such that they can interact        so as to cause a phase separation,    -   the two liquids, at the connecting location, are converted into        a two-phase flow (hereinafter referred to as droplet chain) of        alternating volume fractions of the first and the second liquid        by formation of stable or metastable non-equilibrium phases in        particular due to surface properties (e. g. structure, deposits,        films), geometries and/or flow velocities, and this two-phase        flow passes through a transmission channel extending from the        first connecting location of the first channel with the second        channel, and    -   the two-phase flow, after its transmission and at another        connecting location of the transmission channel with a third and        a fourth channel, is spontaneously divided, in particular due to        surface properties (e. g. structure, deposits, films), geometry        and/or flow velocities, into the third and the fourth channels        as the liquid flows that existed prior to the separation.

Thus, according to the invention, a heterogeneous liquid flow is dividedinto portions by introducing individual volume fractions of a secondliquid, which may be but need not be homogeneous, between successivevolume fractions of the heterogeneous liquid. While these volumefractions of the heterogeneous liquid, lying close to one another, wouldmix without this measure being taken, separation of the individualvolume fractions allows these volume fractions, and thus theheterogeneous liquid, to be transported over large distances without anymixing occurring. Treatment of the volume fraction requires separationof the liquid flow of alternating volume fractions of the two liquids sothat again two flows are produced one of which being the heterogeneousstarting liquid and the other being the second liquid. Since bothprocesses can take place autonomously by self-organization of thephases, an overall solution for the mixing-free transport of aheterogeneous liquid without employment of expensive fractionatingapparatus can be achieved.

The combination of the two liquid flows as well as the separation of theoverall flow into the two liquid flows occur at connecting locations atwhich the respective channels meet each other. At these connectinglocations chambers may be formed.

Preferred embodiments of the invention are stated in the subclaims.

Fundamentals of the Solution

The following subitems 1-3 describe an apparatus for mixing-freetransport of heterogeneous liquids in capillaries or microchannels withcharacteristic cross-section/structure sizes (width, depth) between 1 μmand 5 mm rendered possible by an autonomous “inline” formation andseparation of alternating droplet chains with the aid of a second liquidwhich is immiscible with the starting solution.

1. Defined Droplet Formation by Kinetic Self-Organization andSelf-Production

A second liquid, which is immiscible with the first phase and adapted toexpand to a small extent, is selected. The two liquids should clearlydiffer from each other with regard to their contact angle to the channelwall. The heterogeneous components of the first liquid may not besoluble in the second phase (separation liquid). For an aqueous solutionof biopolymers for example oils are suitable. The formation ofalternating fluid phases (such as oil/lipid and water droplets), with orwithout additives influencing the surface tension, of two or morefluids, which are immiscible with each other, in a continuous flow iscontrolled at the meeting point of two or more capillaries (diameter5-5000 μm) or microchannel structures (structure width 1-5000 μm,structure height 1-5000 μm). The principle of the procedure is shown inthe drawing. The droplet formation is a kinetic procedure which resultsin a non-equilibrium state. The droplet length (mostly larger than thechannel diameter) and thus the droplet volumes (fl-μm) depend on thevolume of the meeting chamber and/or their meeting and/or crossingpoint. Further variables influencing the process are: the selection ofthe second phase (see above), the geometry of the meeting chamber, thewetting properties of the walls/capillary surfaces as well as theselection of various flow rates. The embodiment shows detailed valuesfor two different cases.

2. Autonomous Separation of Alternating Droplet Chains

The autonomous separation of alternating fluid phases (such as oil andwater droplets) under flow conditions is controlled by branchingcapillaries (diameter 5-5000 μm) or microchannel structures (structurewidth 1-5000 μm, structure height 1-5000 μm). Due to the differentwetting properties of the liquids, the aqueous state is discharged inthe drain channel with the hydrophilic surface and the oil phase(separation liquid) is discharged in the drain channel with thehydrophobic surface. In addition, the volume of the separation chamberrelative to the volume of the droplets should be correspondingly small.The two liquids are then neatly separated in the two discharge channels(or capillaries) while flowing at the same velocity as stated underitem 1. Exact conditions for a preferred aspect of the invention arestated in the embodiment.

3. Combination of Items 2 and 1 with Recovery of the Separation PhaseUsed

A complete transfer apparatus requires employment of a reservoir for theseparation-phase liquid (e. g. oil/lipid) and a pump for pumping theliquid from the reservoir. To keep the consumption of theseparation-phase liquid at a low level, the second phase can berecovered. The separation phase is returned, with the aid of the pumpsystem, to the location where the formation process takes place (item 1)such that the systems constitutes a closed circuit.

Improvements and Advantages as Compared with the State of the Art

The transport of heterogeneous molecule samples (in particular in theaqueous phase) split into droplets can take place without the moleculesmixing with each other (prevented by the separation phase, e. g. oildroplets), provided that the molecules are not soluble in the secondphase. By utilization of mobile droplets heterogeneous samples can thusbe separated and transported over larger distances (e. g. severalmeters) without any mixing occurring. In this manner, such separatedsamples can be transported to the location of a following synthesis stepor another analysis. In suspensions of micro-/nanoparticles, too,droplets containing defined quantities of particles (beads) can beformed and fed to the desired reaction and/or analysis locations.Advantage: improvement of the reproducibility of analyses/diagnosticprocedures.

EMBODIMENT

Hereunder the invention is explained in detail with reference to anembodiment shown in the drawing.

Two liquids in the corresponding inlet channels/capillaries meet in ameeting chamber 10 and autonomously form a droplet chain 12. In theexample, a heterogeneous aqueous liquid 14 and an oil/lipid phase 16 areshown as liquids that are immiscible with each other. However, otherimmiscible phases can also be used.

The heterogeneous aqueous liquid 14 is, for example, constituted byindividual sample volumes lying one behind the other which are producedin a sample treatment unit 18 and arranged in series as seen in thedirection of flow. This heterogeneous solution is now to be transportedto a location at a relatively large distance from the production site,where the solution is subjected to further treatment (for example,analysed, fractioned or subjected to other treatment steps). Thislocation is generally shown in the drawing as treatment location 20. Theproblems encountered are that the individual sample volumes can mix witheach other when the heterogeneous aqueous solution is transported over alarger distance, whereas the transport over a relatively short distance(for example, up to 1 cm) does not lead to any disturbing mixing.Therefore, the heterogeneous aqueous liquid is transported from the unit18 via a short channel 22 to the meeting chamber 10. In this meetingchamber 10 the oil/lipid phase 16 is added via a channel 24.

In the meeting chamber 10, due to the mechanisms described above (cf.the aforementioned citations), a spontaneous phase separation of the twoflows occurs and the droplet chain 12 is formed in a channel 26extending from the meeting chamber 10. This droplet chain 12 comprisesvolume fractions alternately lying one behind the other of theheterogeneous aqueous solution 14 and the oil/lipid phase 16. Theseparation takes place in the meeting chamber 10 such that theindividual sample volumes of the aqueous solution 14 are now separatedfrom each other by the oil/lipid volumes. Thus the sample volumes do notmix with each other in the channel 26.

For further treatment of the sample volumes in the unit 20, theindividual sample volumes must be recombined to again form aheterogeneous aqueous solution flow. For this purpose, the droplet chain12 is separated in a separation chamber 28 into which the channel 26leads. The heterogeneous aqueous solution 14, comprised of thesuccessive individual sample volumes, flows out via a first outletchannel 30, while the oil/lipid phase 16 flows out via a second outletchannel 32. This second outlet channel 32 can be connected via a pump 34with the first channel 24 to form a circulation system for the oil/lipidphase. In this circulation system, a reservoir 36 can be arranged. Thepump 34 is required on the one hand to allow substance circulation, andon the other hand to control the transport velocity.

Droplet Formation (in the Meeting Chamber 10)

The formation of very small droplets is achieved by the combination oflow flowing velocities (with magnitude of <1 μl/min.) and small volumesat the crossing point (in the magnitude of some nl). At higher flowrates (in the magnitude of >0.05 ml/min.) a “nozzle effect” is utilizedwhich occurs when oil/lipid is introduced into the flow of an aqueousphase.

(i) Low Flow Rates:

In a T-section (wall material untreated silicon; channel width 300 μm;channel depth 50 μm, volume of the meeting chamber 5 nl) water and oilare combined as shown in the drawing. If the meeting volume is filledwith one of the two liquids and then identical flow rates of the twoliquids are set, water and oil alternately flow into the meeting volume,provided that the flow rates do not exceed 1 μm/min. In the dischargechannel by this means chains of alternating droplets of the two liquidswith a droplet volume of 100 nl can be observed. If the volume of thedroplets formed is scaled to the volume of the meeting chamber, thechannel width of a T-section (channel depth again 50 μm), in which thedroplets of a volume of 500 pl are formed, can be reduced toapproximately 5 μm.

(ii) High Flow Rates:

Polyethylene tubes (inner diameter 400 μm) are connected via a siliconeadapter tube (hydrophobic, inner diameter 500 μm) to a Y-section(hydrophilic wall material, e. g. glass; line diameter 200 μm; length ofline as from crossing point 8 mm). Via two of the tubes water and oilare transported into the Y-section. At a constant flow rate of the waterof 0.05 ml/min., oil flows at flow rates of more than 0.2 ml/min.through the meeting chamber without contacting the hydrophilic glasswalls of the Y-section. The water flows between the oil and the glasswall into the discharge line. Formation of the droplets takes place atthe transition between the glass Y-section and the silicone tube due toa “nozzle effect” and is promoted by the change in the wettingproperties of the walls. The water droplets have a volume of 90 nl, theoil droplets have a volume of 300 μm at a flow rate of 0.2 ml/min.

Separation (in the Separation Chamber 28)

Into a T-section (one of the arms having hydrophilic, the opposite armhaving hydrophobic walls; channel width 10 μm, channel depth 50 μm;volume of the meeting chamber 5 pl) a chain of alternating droplets ofwater and oil (droplet volume 1 nl) is transported at flow rates notexceeding 0.1 μm/min. The chain of alternating droplets is separatedinto an oil flow and a water flow which flow out through the hydrophobicline and the hydrophilic line, respectively.

Transport System

The two system components (droplet formation/separation) can becombined, as shown in the drawing, for the purpose of transportingheterogeneous liquids in a closed-circuit system. In many analysisprocesses (e. g. chromatography) samples are separated and heterogeneoussolutions are produced such as are also produced during kineticprocesses in flow reactors. The existing concentration gradients at theoutput of the measuring instrument and/or the reactor can be separatedinto small microsample volumes with the aid of the described process fordroplet formation and then transported in a mixing-free manner over longdistances to locations of further treatment. There or after thetreatment step, the droplet chains can be recombined and/or separatedinto their phases.

1. Method for mixing-free transport of a heterogeneous liquid inmicrochannels, wherein in the method a first heterogeneous liquid (14)flows through a first channel (22) and, at a first connecting location(meeting chamber 10), meets a second (24) channel carrying a secondliquid (16), wherein the two liquids (14,16) are selected such that theycooperate to cause a phase separation, the first and second liquids(14,16), at the connecting location (meeting chamber 10), are convertedinto a two-phase flow (droplet chain 12) of alternating volume fractionsof the first and second liquids (14,16) by formation of non-equilibriumphases in particular due to surface structures, deposits or films,geometries and/or flow velocities, and this two-phase flow (dropletchain 12) passes through a transmission channel (26) extending from theconnecting location (meeting chamber 10) of the first channel (14) withthe second channel (24), and the two-phase flow (droplet chain 12),after this transmission and at another connecting location (separationchamber 28) of the transmission channel (26) with a third and a fourthchannel (30,32), is spontaneously divided, in particular due to surfacestructures, deposits or films, geometry and/or flow velocities, into thethird and fourth channels (30,32) as the liquid flows that existed priorto the separation.
 2. Method according to claim 1, characterized in thatat least one of the heterogeneous liquids comprises microparticles, suchas beads.
 3. The method of claim 1 wherein the method is applied inchromatography, in particular for protein purification or other(chemical) separation processes.
 4. The method of claim 1, wherein themethod is applied for transporting heterogeneous microparticles, inparticular beads, distributed in at least one of the liquids.
 5. Themethod of claim 2, wherein the method is applied in chromatography, inparticular for protein purification or other (chemical) separationprocesses.
 6. The method of claim 2, wherein the method is applied fortransporting heterogeneous microparticles, in particular beads,distributed in at least one of the liquids.